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Biomedical subjects

H Breer

Publications and source records attributed to H Breer.

At least 109 records · Page 6Linked to original sources
PubMed1993

Second messenger signalling in olfaction.

Odorous molecules are recognized by specific receptor proteins located in the ciliary membrane of olfactory receptor neurons. These receptors have been identified using molecular cloning--they are members of the seven-transmembrane-domain G protein-coupled receptor superfamily. Specific receptor subtypes are expressed in subsets of olfactory neurons spatially segregated within certain areas of the olfactory epithelium. Interaction of odorants with receptors initiates the primary reaction of olfactory signalling. Intracellular reaction cascades are activated via specific G proteins, leading to a rapid and transient rise in second messenger levels; odorous compounds elicit mutually exclusive cAMP or inositol 1,4,5-trisphosphate responses. Odorant-induced second messenger signalling is terminated via kinase-mediated negative feedback loops uncoupling the reaction cascades by phosphorylation of receptor proteins. Strong odour stimuli elicit a delayed response of another messenger system, the nitric oxide/cGMP cascade. cGMP may control some adaptive reactions in olfactory receptor neurons.

Animals↗
PubMed1993

Implications of the NO/cGMP system for olfaction.

The rapid and transient generation of pulses of either cAMP or IP3 is considered the primary reaction in olfactory signal transduction. There is some recent evidence suggesting that another second messenger system, involving cGMP, may also play an important role in olfactory signalling. High doses of odorant elicit a delayed and sustained elevation of cGMP levels due to the operation of the nitric oxide (NO) synthase/guanylyl cyclase system. The interplay of NO and cGMP is supposed to trigger molecular mechanisms, including adaptation processes, which enable the olfactory neuroepithelium to cope with strong stimuli. Furthermore, a characteristic pattern of staining for NO synthase in the olfactory bulb suggests that the NO/cGMP system may also participate in neuronal processing of the sensory input.

Animals↗
PubMed1993

Analysis of rapid calcium signals in synaptosomes.

A combination of the stopped-flow technology with dual channel spectrofluorometry of Ca(2+)-indicators was utilized for the measurement of rapid Ca(2+)-signals in rat cerebral cortical synaptosomes evoked by K(+)-depolarization. There was no observable contribution of Ca(2+)-ions from intracellular stores to the rise in [Ca2+]i. The kinetics of the fast increase in intracellular Ca2+ concentration was analysed in relation to the depolarization strength. The maximal increase in [Ca2+]i and the time course of Ca(2+)-channel inactivation were determined for depolarizations obtained by different extracellular K(+)-concentrations ([K+]o). An apparent threshold was observed at about 18 mM [K+]o; a maximal Ca(2+)-signal amplitude was estimated at about 40 mM [K+]o. Pharmacological properties of the involved Ca(2+)-channels were determined using selective Ca(2+)-channel blockers (Dihydropyridines, omega-Conotoxin, omega-Agatoxins); the results suggest that a P-type voltage-dependent Ca(2+)-channel is the relevant channel type, generating the evoked Ca(2+)-signals in rat cerebral cortical synaptosomes.

Animals↗
PubMed1993

Arrestin-subtypes in insect antennae.

Arrestin is supposed to be involved in uncoupling receptor-mediated second messenger cascades. Clones encoding proteins homologous to arrestin have been isolated from antennal cDNA libraries of Locusta migratoria and Heliothis virescens. Based on the size and several characteristic motifs, the two proteins are considered as members of different arrestin subfamilies. One of the subtypes, which has also been found in Drosophila, lacks the regulatory acidic C-terminal. The putative site of interaction with phosphorylated receptors, a cationic region in the primary structure, is conserved in all identified arrestins from locust to human.

Amino Acid Sequence↗
PubMed1993

Odorant binding proteins of Heliothis virescens.

cDNA clones coding for three different binding proteins were isolated from an antennal library of Heliothis virescens. The deduced amino acid sequences showed only moderate homology to each other but shared several common structural features. Based on a comparison with the predicted primary structures of antennal binding proteins from different moth species, one of the clones (Hel-1) was found to encode a pheromone binding protein, whereas the two others (Hel-10 and -11) encode general odorant binding proteins.

Amino Acid Sequence↗
PubMed1993

Neurotrophin induced second messenger responses in rat brain synaptosomes.

Neurotrophins (NGF, BDNF) elicit significant elevation of cAMP as well as IP3-concentrations in either membrane preparations or intact isolated nerve endings from rat brain hippocampus. The induced second messenger responses in membrane preparations were detectable in the subsecond time range. The IP3-level reached a maximum after a few hundred ms; whereas the cAMP-level continued to rise even after several seconds. The IP3-response but not the cAMP-signal was inhibited in the presence of the tyrosine kinase blocker K252a, suggesting that the two second messenger cascades were triggered via different mechanisms. The results suggest that neurotrophins may induce short-term effects in nerve terminals via second messenger pathways.

Animals↗
PubMed1993

Expression of a mucociliary-specific epitope in human olfactory epithelium.

An olfactory ciliary-specific epitope was localized immunohistochemically in the mucociliary complex of human olfactory epithelium of 12 subjects ranging in age from 16 weeks of gestation to 85 years, including 3 with Alzheimer's disease. Immunoreactivity for olfactory marker protein (OMP) was used to identify olfactory epithelium; OMP immunoreactivity in olfactory receptor neurons in a 16-week old fetus is the earliest time point at which OMP expression has been detected in human gestation. The results suggest a close coupling between the expression of ciliary molecules associated with odorant transduction and the functional maturation of olfactory receptor neurons.

Adult↗
PubMed1993

Rapid kinetic measurements of second messenger formation in olfactory cilia from channel catfish.

The effect of stimulating olfactory cilia from the channel catfish (Ictalurus punctatus) with odorant amino acids on the formation of adenosine 3',5'-cyclic monophosphate, guanosine 3',5'-cyclic monophosphate, and inositol 1,4,5-trisphosphate (IP3) was studied in the subsecond time scale using a quenched flow technique. L-Alanine (L-Ala) and L-cysteine (L-Cys) (100 microM) elicited a transient elevation in IP3 levels that peaked at 25 ms. In contrast, even at high concentration a mixture of odorant amino acids (1 mM L-Ala, L-Cys, L-norleucine, L-glutamate, L-proline, and L-arginine) did not elicit a change in cAMP levels in this time scale and caused only relatively slow and minor increases in cGMP. The dose-response relationship for the IP3 response of L-Cys and L-Ala in the range from 10 nM to 1 mM is consistent with previous electrophysiological and ligand binding experiments. Odorant amino acid-stimulated IP3 formation was GTP dependent and was inhibited by guanosine 5'-O-(2-thiodiphosphate), suggesting that the response was G protein mediated. These results are consistent with a mediatory role for IP3 in amino acid olfactory signal transduction in catfish.

Alanine↗
PubMed1993

Generation of monoclonal antibodies detecting specific epitopes in locust antennae.

1. Following a tissue-specific screening paradigm, monoclonal antibodies have been generated that interact with distinct subpopulations of cells in locust antennae. 2. Antigens were identified as high molecular weight components. 3. Immunoreactivity was not detectable during embryonic development, but rapidly appeared within a few hours of hatching. 4. The time course of antigen expression in antennal cells could be followed in situ as well as in vitro. 5. Expression of monoclonal antibody B14/6D2-like immunoreactivity was prevented by blocking protein synthesis with cycloheximide.

Animals↗
PubMed1993

Towards an identification of odorant receptors.

The molecular nature and diversity of receptive sites for odorous molecules is a central unanswered issue in olfaction. Based on the enormous resolving power of the olfactory system, which enables the stereospecific discrimination of numerous compounds at low concentrations specific receptor proteins have been proposed. Due to the central role of G-proteins in olfactory signalling it has been predicted that odorant receptors might be members of the superfamily of receptor proteins with seven transmembrane domains. Upon application of degenerated oligonucleotides and the PCR-technology a number of putative odorant receptors have been cloned and sequenced. In situ hybridisation studies using receptor-specific probes have been performed and olfactory neurons specifically expressing a particular receptor subtype are topologically identified in the nasal epithelium of rats. Clones of interest are now being expressed in heterologous systems in order to demonstrate functional activity of these putative receptor proteins and to match defined odorants to identified receptors.

Amino Acid Sequence↗
PubMed1992

Odor-induced phosphorylation of olfactory cilia proteins.

Stimulation of isolated rat olfactory cilia in the presence of [gamma-32P]ATP leads to a significantly enhanced incorporation of [32P]phosphate. Depending on the type of odorants applied, the induced phosphorylation is completely blocked by specific inhibitors of either protein kinase A or protein kinase C. Time-course experiments indicate that the odor-induced modification of ciliary proteins is transient; the intensity of labeling decayed over time (1-10 sec). Separation of ciliary proteins by SDS/polyacrylamide gel electrophoresis followed by autoradiography demonstrated that upon stimulation with lilial, a single polypeptide (50,000 Da) was phosphorylated; the size of the modified protein is in line with the hypothesis that odorant receptors are phosphorylated subsequent to activation by specific odors.

Animals↗
PubMed1992

Termination of second messenger signaling in olfaction.

By using isolated rat olfactory cilia and a fast kinetics methodology, it has been demonstrated that odorant-induced second messenger signaling in the millisecond time range is terminated via phosphorylation reactions catalyzed by specific protein kinases. The cyclic adenosine nucleotide pathway is turned off by kinase A activity, whereas the inositol trisphosphate cascade is terminated by kinase C. The data support the concept that desensitization of odorant responses involves phosphorylation of key elements in the transduction cascade.

1-Methyl-3-isobutylxanthine↗
PubMed1992

Expression of a pheromone-binding protein in insect cells using a baculovirus vector.

A cDNA encoding a pheromone-binding protein from the male silkmoth Antheraea pernyi has been integrated into the genome of the Autographa californica multiple nuclear polyhydrosis virus such that the transcription was under the control of the strong polyhedrin promoter. Recombinant pheromone-binding protein was expressed in a baculovirus-infected insect cell line (Sf9) and secreted from the cells into the culture medium. Using a two-step protocol, recombinant pheromone-binding protein has been isolated and purified to homogeneity. Pheromone binding of recombinant protein has been demonstrated using a tritiated analog of (E,Z)-6,11-hexadecadienyl acetate.

Affinity Labels↗
PubMed1992

Regulation of hemicholinium binding sites in isolated nerve terminals.

High-affinity uptake of choline, the rate-limiting, regulatory step for the synthesis of acetylcholine is regulated via presynaptic auto- and heteroreceptors. Binding studies using tritiated hemicholinium-3 ([3H]HCh-3) as the specific ligand for the choline carrier revealed that the number of hemicholinium binding sites in nerve terminals isolated from insect brain changes corresponding to the activity of synaptosomal kinase A and kinase C. Activation of kinase A apparently increases the total number of hemicholinium binding sites by recruiting additional occult carriers, whereas the effect of kinase C activity is most appropriately explained by preventing a down-regulation of carrier proteins. The kinase-mediated regulation of choline transporters is obviously due to a phosphorylation of the carrier protein itself.

Animals↗
PubMed1992

Rapid kinetics of depolarization-induced changes in intrasynaptosomal calcium concentrations.

The rapid kinetics of depolarization-evoked calcium influxes in isolated nerve terminals from rat cortex were monitored by stopped-flow spectrofluorimetry using specific indicators (Fluo-3, Indo-1). A very rapid increase in the intrasynaptosomal Ca(2+)-level was detected within the subsecond time range after depolarizing synaptosomes by mixing with physiological saline containing elevated K(+)-concentrations. About 15 mM [K+]o was determined as threshold concentration for inducing Ca(2+)-influx, which increased with higher concentration and saturated at [K+]o-concentrations of about 40 mM [K+]o.

Aniline Compounds↗
PubMed1992

Second messenger signalling in olfaction.

The primary reactions of the chemo-electrical signal transduction pathway in olfactory receptor neurons are mediated by two alternative second messengers, cAMP and inositol 1,4,5-trisphosphate. The rapid and transient intracellular signalling is terminated by the action of negative-feedback loops which uncouple the reaction cascades (desensitization). Recent evidence suggests that secondary reactions in olfaction (adaptation) may also be controlled by second messengers.

Animals↗
PubMed1992

Nitric oxide mediated formation of cyclic GMP in the olfactory system.

Olfactory cilia preparation from rats contain considerable activity of soluble guanylate cyclase as indicated by the formation of cyclic GMP (cGMP) upon application of nitroprusside, a nitric oxide generating agent. Stimulation of olfactory cilia with high doses of odorants elicited a delayed and sustained elevation of the cGMP-concentration. The odorant-induced cGMP-response was abolished by L-NG-nitro-arginine, a selective inhibitor of nitric oxide synthesis, as well as by haemoglobin which efficiently binds and inactivates nitric oxide. These observations suggest that the NO/cGMP cascade may plan an important role in signal processing of the olfactory system.

Animals↗